To assess the microbiological status of a biofiltration system used to treat borehole water, filtermatrix samples were analyzed after staining for the presence of active biofilms using confocal laser scanningmicroscopy (CLSM). CLSM revealed the presence of biofilms on the filter matrix with actively metabolizingmicrobial cells present. Thereafter, heterotrophs,manganese oxidizing bacteria (MOB) and iron oxidizing bacteria(IOB) present in the biofilms were quantified. For heterotrophs a count of 2.9×107 cfu/g was established usingR2A agar while counts for presumptive MOB and IOB were established as 2.4×107 cfu/g and 3.1×107 cfu/grespectively. In addition, a clone library was established using DNAextracted from a pooled filter matrix sampleto assess the diversity of bacteria present within the biofilter matrix. A total of 100 randomly selected cloneswere separated into 14 unique operational taxonomic unit (OTU’s) based upon restriction patterns of amplifiedpartial 16S rRNA genes. Overall, 38% of the clones were assigned to the phylum Proteobacteria, 13% to thephylum Actinobacteria, 24% to the phylum Firmicutes, 21% to the phylum Nitrospirae and 4% to the phylumVerrucomicrobia.